Method for immobilizing protein by oriented affinity of conducting polymer modified vector

A conductive polymer and protein technology, applied to biochemical equipment and methods, and enzymes immobilized on or in inorganic carriers, can solve the problems of decreased activity of immobilized proteins, achieve biological activity retention, shorten distance, and improve active effect

Inactive Publication Date: 2016-11-16
ZHEJIANG SCI-TECH UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the immobilized protein structure obtained by the traditional protein immobilization method is that the protein is connected to the

Method used

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  • Method for immobilizing protein by oriented affinity of conducting polymer modified vector
  • Method for immobilizing protein by oriented affinity of conducting polymer modified vector
  • Method for immobilizing protein by oriented affinity of conducting polymer modified vector

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0034] (1) 10 mg of graphene oxide (GO) was ultrasonically dispersed in 10 mL of pure water for 30 min to prepare a 1 mg / mL GO suspension. Weigh 500mg NaOH and 500mg ClCH 2 COONa was added into the GO suspension, ultrasonically reacted for 2 h, and the obtained product was neutralized with dilute HCl. Repeat washing with deionized water and centrifugation (10000rpm, 10min) until the product is well dispersed in deionized water. Finally, use a dialysis bag in deionized water, dialyze (molecular weight cut off: 8000-10000) for 48 hours to remove ions, and dilute the obtained GO-COOH suspension to 10 mL.

[0035] (2) Weigh 100mg (0.15mol) of Hemin into a three-necked flask, add 40mL of THF, stir to dissolve; raise the temperature to 70°C, and gradually add thionyl chloride (0.6mol) dropwise; after the dropwise addition, raise the temperature to 110°C , continue to react for 1h. The unreacted thionyl chloride and the solvent in the above reaction solution were distilled off to ...

Embodiment 2

[0041] Detection of hydrogen peroxide (H 2 o 2 ) The current response relationship of the substrate

[0042] (1) Prepare 78 μM H 2 o 2 solution;

[0043] (2) With the above-mentioned enzyme sensor as the catalytic electrode, the substrate is 78 μM H 2 o 2 solution; the measuring voltage is 0-0.5V, and the scanning rate is 50mV / s; the cyclic voltammetry curve (CV) of the biocatalytic system is tested.

[0044] exist figure 1 Among them, a is the unreduced modified enzyme, b is the reduction degree of 0.25, c is the reduction degree of 0.5, d is the CV curve of the reduction degree of 1 and e the reduction degree of 2. The measured catalytic current increases with increasing reduction degree. The electrochemical activity of the modified enzyme is much larger than that of the modified one, which is about 2 times that of the unmodified enzyme.

[0045] figure 2 And in Table 1, with -0.37V as the measurement voltage, the I-t curve of the HRP response was detected and obtai...

Embodiment 3

[0050] (1) 15 mg of carbon nanotubes were ultrasonically dispersed in 10 mL of pure water for 30 min to prepare a carbon nanotube suspension. Weigh 300mgNaOH and 300mgClCH 2 COONa was added to the carbon nanotube suspension, subjected to ultrasonic reaction for 2 h, and the obtained product was neutralized with dilute HCl. Washing with deionized water and centrifugation (10000 rpm, 10 min) were repeated until the product was well dispersed in deionized water to obtain a suspension of carboxylated carbon nanotubes.

[0051] (2) Weigh 100mg (0.15mol) of Hemin into a three-necked flask, add 40mL of THF, stir to dissolve; raise the temperature to 70°C, and gradually add thionyl chloride (0.6mol) dropwise; after the dropwise addition, raise the temperature to 110°C , continue to react for 1h. The unreacted thionyl chloride and the solvent in the above reaction solution were distilled off to obtain the acid chlorided Hemin.

[0052] Under cooling in an ice-water bath, weigh 0.5 g...

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Abstract

The invention discloses a method for immobilizing protein by oriented affinity of a conducting polymer modified vector. According to the method disclosed by the invention, the vector is modified by a conducting polymer, and oriented immobilized protein is constructed on the basis of biological affinity between prosthetic group and apoprotein. The method disclosed by the invention has main advantages that the conductive vector is constructed by virtue of the conducting polymer; in addition, the protein is highly oriented, which is more conducive to the contact between an active center of the protein and a reaction substrate; and the protein, which is immobilized on the basis of the oriented affinity, is obviously greater than protein, which is immobilized by virtue of a normal adsorption method, in activity; therefore, a feasible method is provided for further constructing a high-efficiency and low-cost bio-catalytic system.

Description

technical field [0001] The invention relates to a method for immobilizing proteins with directional affinity effect by modifying carriers with conductive polymers. Background technique [0002] Protein is the material basis of life, an organic macromolecule, the basic organic matter that constitutes a cell, and the main bearer of life activities. Without protein there is no life. Amino acids are the basic building blocks of proteins. It is a substance closely related to life and various forms of life activities. Every cell and every major part of the body has proteins involved. Protein accounts for 16%-20% of the body weight, that is, a 60kg adult has about 9.6-12kg of protein in his body. There are many types of proteins in the human body, with different properties and functions, but they are all composed of more than 20 amino acids (Amino acid) in different proportions, and are constantly metabolized and renewed in the body. [0003] The immobilization of biologically...

Claims

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Application Information

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IPC IPC(8): C12N11/14
CPCC12N9/0065C12N11/14C12Y111/01005C12Y111/01006C12Y111/01007
Inventor 叶鹏胡玲玲许永娟
Owner ZHEJIANG SCI-TECH UNIV
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